This invention relates to affinity chromatography. More particularly, the invention concerns the use of affinity chromatography matrices comprising a support, a bifunctional spacer and a compound (ligand) having a high and specific affinity for the compound sought to be isolated and/or purified. Even more particular, the invention relates to affinity chromatography matrices useful for purifying active interleukin-1.beta. converting enzymes (ICE).
The literature is replete with articles, monographs, and books on the subject of affinity chromatography, including such topics as affinity chromatography supports, crosslinking members, ligands and their preparation and use. A sampling of those references includes:
Affinity chromatography: general methods. Methods Enzymol. 1990, 182 (Guide Protein Purif.), 357-71; PA0 Novel affinity-based processes for protein purification. J. Ferment. Bioeng. 1990, 70(3), 199-209; PA0 Applications of preparative high-performance liquid chromatography to the separation and purification of peptides and proteins. J. Chromatogr. 1989, 492, 431-69; PA0 Large-scale purification of enzymes. Appl. Microbiol. Res., Ciba Found. Symp. 1985, 111(Enzymes Org. Synth.), 40-56 (Eng).; PA0 Purification of enzymes by heparin-Sepharose affinity chromatography. J. Chromatogr. 1980, 184(3), 335-45; PA0 Principles of multi-enzyme purification by affinity chromatography. Enzyme Eng. 1978, 4, 441-2; PA0 Purification of enzymes and other biologically active proteins by chromatography on low-molecular-weight substances immobilized on solid supports. Postepy Biochem. 1977, 23(1), 113 27; PA0 General ligand affinity chromatography in enzyme purification. Ligands, affinity chromatography, enzyme purification. J. Macromol. Sci., Chem. 1976, A10(1-2), 15-52; PA0 Affinity purification of enzymes. Chem. Technol. 1975, 5(9), 564-71; PA0 Chromatography, affinity. Encycl. Polym. Sci. Eng. 1985, 3, 531-48; PA0 Bioaffinity chromatography. Pract. High Perform. Liq. Chromatogr. 1976, 193-206; PA0 Affinity chromatography of plasma proteins--an overview. Proc. Int. Workshop Technol. Protein Sep. Improv. Blood Plasma Fractionation, 1977, 422-35; PA0 Affinity chromatography of enzymes. Affinity Chromatogr., Proc. Int. Symp. 1977 (Pub. 1978), 25-38; PA0 Protein immobilization and affinity chromatography. Biotechnol. Appl. Proteins Enzymes. Pap. Conf. 1976 (Pub. 1977), 83-102; PA0 Use of affinity chromatography in protein structure studies. Pept., Proc. Eur. Pept. Symp., 11th 1971 (Pub. 1973), 203-22; PA0 Affinity chromatography of enzymes. Fed. Eur. Biochem. Soc. Meet., [Proc] 1974, 30; PA0 Support materials for immobilized enzymes and affinity chromatography. Bonded Stationary Phases Chromatogr. 1974, 93-112; PA0 Supports for immobilized enzymes and affinity chromatography. Chem. Technol. 1974, 4(11), 694-700; PA0 Affinity chromatography. Enzyme-inhibitor systems. Methodol. Develop Biochem. 1973, 2, 109-12; PA0 Affinity chromatography. Specific separation of proteins. Chromatographia 1971, 4(12), 578-87; PA0 Affinity chromatography. A Practical approach, IRL Press Limited, Oxford England (1985); and PA0 Methods in Enzymology, Vol. 34: Affinity Techniques. Enzyme Purification: 1974. 810 pp.
A wide variety of affinity chromatography supports (with and without spacers) and support/ligands (with and without spacers) are disclosed in the SIGMA catalog, SIGMA CHEMICAL Co., St. Louis, Mo.
Interleukin-1.beta. converting enzyme (ICE) has been identified as the enzyme responsible for converting precursor interleukin-1.beta. (IL-1.beta.) to biologically active IL-1.beta.. Accordingly, interleukin-1.beta. converting enzyme is useful in the diagnosis of IL-1 mediated diseases or in enhancing the production of IL-1.
Mammalian interleukin-1 (IL-1) is an immunoregulatory protein secreted by cell types as part of the inflammatory response. The primary cell type responsible for IL-1 production is the peripheral blood monocyte. Other cell types have also been described as releasing or containing IL-1 or IL-1 like molecules. These include epithelial cells (Luger, et al., J. Immunol. 127: 1493-1498 (1981), Le et al., J. Immunol. 138: 2520-2526 (1987) and Lovett and larsen, J. Clin. Invest. 82: 115-122 (1988), connective tissue cells (Ollivierre et al., Biochem. Biophys. Res. Comm. 141: 904-911 (1986), Le et al, J. Immunol. 138: 2520-2526 (1987), cells of neuronal origin (Giulian et al., J. Esp. Med. 164: 594-604 (1986) and leukocytes (Pistoia et al., J. Immunol. 136: 1688-1692 (1986), Acres et al., Mol. Immuno. 24: 479-485 (1987), Acres et al., J. Immunol. 138: 2132-2136 (1987) and Lindenmann et al., J. Immunol 140: 837-839 (1988).
Biologically active IL-1 exists in two distinct forms, IL-1.alpha. with an isoelectric point of about pI 5.2 and IL-1.beta. with an isoelectric point of about 7.0 with both forms having a molecular mass of about 17,500 (Bayne et al., J. Esp. Med. 163: 1267-1280 (1986) and Schmidt, J. Esp. Med. 160: 772 (1984). The polypeptides appear evolutionarily conserved, showing about 27-33% homology at the amino acid level (Clark et al., Nucleic Acids Res. 14: 7897-7914 (1986).
Mammalian IL-1.beta. is synthesized as a cell associated precursor polypeptide with a molecular mass of about 31.4 kDa (Limjuco et al., Proc. Natl. Acad. Sci USA 83: 3972-3976 (1986). Precursor IL-1.beta. is unable to bind to IL-1 receptors and is biologically inactive (Mosley et al., J. Biol. Chem. 262: 2941-2944 (1987). Biological activity appears dependent upon some form of proteolytic processing which results in the conversion of the precursor 31.5 kDa form to the mature 17.5 kDa form. Evidence is growing that by inhibiting the conversion of precursor IL-1.beta. to mature IL-1.beta., one can effectively inhibit the activity of interleukin-1.
Mammalian cells capable of producing IL-1.beta. include, but are not limited to, karatinocytes, endothelial cells, mesangial cells, thymic epithelial cells, dermal fibroblasts, chondrocytes, astrocytes, glioma cells, mononuclear phagocytes, granulocytes, T and B lymphocytes and NK cells.
As discussed by J. J. Oppenheim, et al. Immunology Today, vol. 7(2): 45-56 (1986), the activities of interleukin-1 are many. It has been observed that catabolin, a factor that promotes degradation of cartilage matrix, also exhibited the thymocyte comitogenic activities of IL-1 and stimulates chondrocytes to release collagenase neutral proteases and plasminogen activator. In addition, a plasma factor termed proteolysis inducing factor stimulates muscle cells to produce prostaglandins which in turn leads to proteolysis, the release of amino acids and, in the long run, muscle wasting, and appears to represent a fragment of IL-1 with fever-inducing, acute phase response and thymocyte co-mitogenic activities.
IL-1 has multiple effects on cells involved in inflammation and wound healing. Subcutaneous injection of IL-1 leads to margination of neutrophils and maximal extravascular infiltration of the polymorphonuclear leukocytes (PMN). In vitro studies reveal IL-1 to be a chemotactic attractant for PMN to activate PMN to metabolize glucose more rapidly to reduce nitroblue tetrazolium and to release their lysozomal enzymes. Endothelial cells are stimulated to proliferate by IL-1 to produce thromboxane, to become more adhesive and to release procoagulant activity. IL-1 also enhances collagen type IV production by epidermal cells, induces osteoblast proliferation and alkaline phosphatase production and stimulates osteoclasts to resorb bone. Even macrophages have been reported to be chemotactically attracted to IL-1 to produce prostaglandins in response to IL-1 and to exhibit a more prolonged and active tumoricidal state.
IL-1 is also a potent bone resorptive agent capable upon infusion into mice of causing hypercaleemia and increase in bone resorptive surface as revealed by histomorphometry Sabatini, M. et al., PNAS 85: 5235-5239, 1988.
Accordingly, IL-1 has been implicated in infectious diseases where active infection exists at any body site, such as meningitis and salpingitis; complications of infections including septic shock, disseminated intravascular coagulation, and/or adult respiratory distress syndrome; acute or chronic inflammation due to antigen, antibody, and/or complement deposition; inflammatory conditions including arthritis, cholangitis, colitis, encephalitis, endocarditis, glomerulonephritis, hepatitis, myocarditis, pancreatitis, pericarditis, reperfusion injury and vasculitis. Immune-based diseases which may be responsive to ICE inhibitors of Formula I include but are not limited to conditions involving T-cells and/or macrophages such as acute and delayed hypersensitivity, graft rejection, and graft-versus-host-disease; auto-immune diseases including Type I diabetes mellitus and multiple sclerosis. IL-1 has also been implicated in the treatment of bone and cartilage resorption as well as diseases resulting in excessive deposition of extracellular matrix. Such diseases include periodonate diseases interstitial pulmonary fibrosis, cirrhosis, systemic sclerosis, and keloid formation.